Mechanisms of PM10 Disruption of the Nrf2 Pathway in Cornea.

We have previously shown that PM10 exposure causes oxidative stress and reduces Nrf2 protein levels, and SKQ1 pre-treatment protects against this damage in human corneal epithelial cells (HCE-2). The current study focuses on uncovering the mechanisms underlying acute PM10 toxicity and SKQ1-mediated protection. HCE-2 were pre-treated with SKQ1 and then exposed to 100 µg/mL PM10. Cell viability, oxidative stress markers, programmed cell death, DNA damage, senescence markers, and pro-inflammatory cytokines were analyzed. Nrf2 cellular location and its transcriptional activity were determined. Effects of the Nrf2 inhibitor ML385 were similarly evaluated. Data showed that PM10 decreased cell viability, Nrf2 transcriptional activity, and mRNA levels of antioxidant enzymes, but increased p-PI3K, p-NFκB, COX-2, and iNOS proteins levels. Additionally, PM10 exposure significantly increased DNA damage, phosphor-p53, p16 and p21 protein levels, and ß-galactosidase (ß-gal) staining, which confirmed the senescence. SKQ1 pre-treatment reversed these effects. ML385 lowered the Nrf2 protein levels and mRNA levels of its downstream targets. ML385 also abrogated the protective effects of SKQ1 against PM10 toxicity by preventing the restoration of cell viability and reduced oxidative stress. In conclusion, PM10 induces inflammation, reduces Nrf2 transcriptional activity, and causes DNA damage, leading to a senescence-like phenotype, which is prevented by SKQ1.

Cystic Echinococcosis of the Liver: Correlation Between Intra-Operative Ultrasound and Pre-Operative Imaging.

Background: Imaging plays an essential role in the management of hepatic hydatid cysts (HCE). The objective of our study was to determine the correlation between pre-operative ultrasound, computed tomography (CT), and intra-operative ultrasound (IOUS) in studying the characteristics and complications of HCE. Patients and Methods: This was a prospective, descriptive, and analytical study conducted in the General Surgery Department of Habib Bourguiba Hospital in Sfax. The study included patients with HCE who underwent conservative surgery between April 2017 and June 2022. Results: We enrolled 49 patients with 94 cysts. At the end of our study, IOUS allowed for better detection of HCE (98.8%) regardless of the number of cysts per patient. IOUS and CT were accurate in studying the location of cysts (κ = 1), whereas pre-operative abdominal ultrasound was less efficient (κ = 0.870). IOUS was the best examination for detecting exocysts (κ = 0.961), studying daughter cysts (κ = 0.823), and exploring vascular relations, but it was less effective (κ = 0.523) in detecting calcifications. Regarding classifications, ultrasound and CT had similar results. However, IOUS was most reliable in differentiating between CE3b and CE4 types (κ = 0.653). Ultrasound, CT, and IOUS were not sensitive in detecting latent HCE suppurations and cystobiliary fistulas. Conclusions: Performing IOUS is essential to prevent recurrences and reduce post-operative morbidity.

Cationized gelatin-sodium alginate polyelectrolyte nanoparticles encapsulating moxifloxacin as an eye drop to treat bacterial keratitis.

A mucoadhesive polyelectrolyte complex (PEC) nanoparticles were developed for ocular moxifloxacin (Mox) delivery in Bacterial Keratitis (BK). Moxifloxacin-loaded G/CG-Alg NPs were prepared by an amalgamation of cationic polymers (gelatin (G)/cationized gelatin (CG)), and anionic polymer (sodium alginate (Alg)) along with Mox respectively. Mox@CG-Alg NPs were characterized for physicochemical parameters such as particle size (DLS technique), morphology (SEM analysis), DSC, XRD, encapsulation efficiency, drug loading, mucoadhesive study (by texture analyzer), mucin turbidity, and viscosity assessment. The NPs uptake and toxicity of the formulation were analyzed in the Human Corneal Epithelial (HCE) cell line and an ocular irritation study was performed on the HET-CAM. The results indicated that the CG-Alg NPs, with optimal size (217.2 ± 4 nm) and polydispersity (0.22 ± 0.05), have shown high cellular uptake in monolayer and spheroids of HCE. The drug-loaded formulation displayed mucoadhesiveness, trans-corneal permeation, and sustained the release of the Mox. The anti-bacterial efficacy studied on planktonic bacteria/biofilms of P. aeruginosa and S. aureus (in vitro) indicated that the Mox@CG-Alg NPs displayed low MIC, higher zone of bacterial growth inhibition, and cell death compared to free Mox. A significant reduction of bacterial load was observed in the BK-induced mouse model.

Analysis of the mucosal chemokines CCL28, CXCL14, and CXCL17 in dry eye disease: An in vitro and clinical investigation.

Mucosal chemokines have antimicrobial properties and play an important role in mucosal immunity. However, little is known about their expression on the ocular surface. This study aimed to analyze the expression of the mucosal chemokines CCL28, CXCL14 and CXCL17 in corneal and conjunctival epithelial cells under in vitro dry eye (DE) conditions, and in conjunctival samples from healthy subjects and DE patients. Human corneal epithelial cells (HCE) and immortalized human conjunctival epithelial cells (IM-HConEpiC) were incubated under hyperosmolar (400-500 mOsM) or inflammatory (TNF-α 25 ng/mL) conditions for 6 h and 24 h to measure CCL28, CXCL14, and CXCL17 gene expression by RT-PCR and their secretion by immunobead-based analysis (CCL28, CXCL14) and ELISA (CXCL17). Additionally, twenty-seven DE patients and 13 healthy subjects were included in this study. DE-related questionnaires (OSDI, mSIDEQ and NRS) evaluated symptomatology. Ocular surface integrity was assessed using vital staining. Tactile sensitivity was measured with Cochet-Bonnet esthesiometer, and mechanic and thermal (heat and cold) sensitivity using Belmonte's non-contact esthesiometer. Subbasal nerve plexus and dendritic cell density were analyzed by in vivo confocal microscopy. Conjunctival cells from participants were collected by impression cytology to measure mucosal chemokines gene expression by RT-PCR. Our results showed that HCE and IM-HConEpiC cells increased CCL28, CXCL14, and CXCL17 secretion under hyperosmolar conditions. The gene expression of CCL28 was significantly upregulated in conjunctival samples from DE patients. CCL28 expression correlated positively with symptomatology, corneal staining, heat sensitivity threshold, and dendritic cell density. CXCL14 expression correlated positively with age, ocular pain, conjunctival staining, tactile sensitivity, and image reflectivity. CXCL17 expression correlated positively with corneal staining. These results suggest that corneal and conjunctival epithelial cells could be a source of CCL28, CXCL14, and CXCL17 on the ocular surface and that CCL28 might be involved in DE pathogenesis.

Protective Effect of Probiotics against Pseudomonas aeruginosa Infection of Human Corneal Epithelial Cells.

Probiotic therapy needs consideration as an alternative strategy to prevent and possibly treat corneal infection. This study aimed to assess the preventive effect of Lactobacillus reuteri and Bifidobacterium longum subsp. infantis on reducing the infection of human corneal epithelial (HCE) cells caused by Pseudomonas aeruginosa. The probiotics' preventive effect against infection was evaluated in cell monolayers pretreated with each probiotic 1 h and 24 h prior to P. aeruginosa challenge followed by 1 h and 24 h of growth in combination. Cell adhesion, cytotoxicity, anti-inflammatory, and antinitrosative activities were evaluated. L. reuteri and B. longum adhered to HCE cells, preserved occludin tight junctions' integrity, and increased mucin production on a SkinEthicTM HCE model. Pretreatment with L. reuteri or B. longum significantly protected HCE cells from infection at 24 h, increasing cell viability at 110% (110.51 ± 5.15; p ≤ 0.05) and 137% (137.55 ± 11.97; p ≤ 0.05), respectively. Each probiotic showed anti-inflammatory and antinitrosative activities, reducing TNF-α level (p ≤ 0.001) and NOx amount (p ≤ 0.001) and reestablishing IL-10 level (p ≤ 0.001). In conclusion, this study demonstrated that L. reuteri and B. longum exert protective effects in the context of corneal infection caused by P. aeruginosa by restoring cell viability and modulating inflammatory cytokine release.

(‒)-Epicatechin reveals amoebicidal activity against Acanthamoeba castellanii by activating the programmed cell death pathway.

BACKGROUND: Acanthamoeba is an opportunistic pathogen that can cause human infections such as granulomatous amebic encephalitis and acanthamoeba keratitis. However, no specific drug to treat the diseases has been developed. Therefore, the discovery or development of novel drugs for treating Acanthamoeba infections is urgently needed. The anti-protozoan activity of (‒)-epicatechin (EC) has been reported, suggesting it is an attractive anti-protozoal drug candidate. In this study, the amoebicidal activity of EC against A. castellanii was assessed and its mechanism of action was unveiled. METHODS: The amoebicidal activity of EC against A. castellanii trophozoites and the cytotoxicity of EC in HCE-2 and C6 cells were determined with cell viability assay. The underlying amoebicidal mechanism of EC against A. castellanii was analyzed by the apoptosis/necrosis assay, TUNEL assay, mitochondrial dysfunction assay, caspase-3 assay, and quantitative reverse transcription polymerase chain reaction. The cysticidal activity of EC was also investigated. RESULTS: EC revealed amoebicidal activity against A. castellanii trophozoites with an IC50 of 37.01 ± 3.96 µM, but was not cytotoxic to HCE-2 or C6 cells. EC induced apoptotic events such as increases in DNA fragmentation and intracellular reactive oxygen species production in A. castellanii. EC also caused mitochondrial dysfunction in the amoebae, as evidenced by the loss of mitochondrial membrane potential and reductions in ATP production. Caspase-3 activity, autophagosome formation, and the expression levels of autophagy-related genes were also increased in EC-treated amoebae. EC led to the partial death of cysts and the inhibition of excystation. CONCLUSION: EC revealed promising amoebicidal activity against A. castellanii trophozoites via programmed cell death events. EC could be a candidate drug or supplemental compound for treating Acanthamoeba infections.

Polymer Nanoparticles with 2-HP-ß-Cyclodextrin for Enhanced Retention of Uptake into HCE-T Cells.

Triamcinolone acetonide (TA), a medium-potency synthetic glucocorticoid, is primarily employed to treat posterior ocular diseases using vitreous injection. This study aimed to design novel ocular nanoformulation drug delivery systems using PLGA carriers to overcome the ocular drug delivery barrier and facilitate effective delivery into the ocular tissues after topical administration. The surface of the PLGA nanodelivery system was made hydrophilic (2-HP-ß-CD) through an emulsified solvent volatilization method, followed by system characterization. The mechanism of cellular uptake across the corneal epithelial cell barrier used rhodamine B (Rh-B) to prepare fluorescent probes for delivery systems. The triamcinolone acetonide (TA)-loaded nanodelivery system was validated by in vitro release behavior, isolated corneal permeability, and in vivo atrial hydrodynamics. The results indicated that the fluorescent probes, viz., the Rh-B-(2-HP-ß-CD)/PLGA NPs and the drug-loaded TA-(2-HP-ß-CD)/PLGA NPs, were within 200 nm in size. Moreover, the system was homogeneous and stable. The in vitro transport mechanism across the epithelial barrier showed that the uptake of nanoparticles was time-dependent and that NPs were actively transported across the epithelial barrier. The in vitro release behavior of the TA-loaded nanodelivery systems revealed that (2-HP-ß-CD)/PLGA nanoparticles could prolong the drug release time to up to three times longer than the suspensions. The isolated corneal permeability demonstrated that TA-(2-HP-ß-CD)/PLGA NPs could extend the precorneal retention time and boost corneal permeability. Thus, they increased the cumulative release per unit area 7.99-fold at 8 h compared to the suspension. The pharmacokinetics within the aqueous humor showed that (2-HP-ß-CD)/PLGA nanoparticles could elevate the bioavailability of the drug, and its Cmax was 51.91 times higher than that of the triamcinolone acetonide aqueous solution. Therefore, (2-HP-ß-CD)/PLGA NPs can potentially elevate transmembrane uptake, promote corneal permeability, and improve the bioavailability of drugs inside the aqueous humor. This study provides a foundation for future research on transocular barrier nanoformulations for non-invasive drug delivery.

Photoresponsive 'chemo-free' phytotherapy: formulation development for the treatment of triple-negative breast cancer.

Aim: The present investigation aimed to develop a chemo-free, nanophytosomal system to treat triple-negative breast cancer (TNBC) via a phyto-photo dual treatment strategy. Method: Size, shape, surface analysis, photoprovoked release profile, photothermal stability, (3-(4, 5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide assay, apoptotic assay, DNA fragmentation, in vitro cellular uptake evaluation, mitochondrial membrane potential and caspase-3 assay, and photodynamic evaluation. Results: Biological experiments using MDA-MB-231 cells displayed dose-dependent synergistic anti-TNBC activity of PhytoS/Houttuynia cordata extract (HCE)/IR780 as compared with Phyto/HCE, PhytoS/IR780 and even more promising under laser treatment. Apoptotic assay and DNA fragmentation analysis also showed enhanced anti-TNBC effects. Investigation found that HCE acts via suppression of mitochondrial membrane potential and inducing caspase-3 activity in cells. Conclusion: Our findings suggested that photo-empowered phytotherapy can be employed effectively and safely against TNBC.

Hemerocallis citrina Baroni ameliorates chronic sleep deprivation-induced cognitive deficits and depressive-like behaviours in mice.

Sleep deprivation (SD) is common during spaceflight. SD is known to cause cognitive deficits and depression, requiring treatment and prevention. Hemerocallis citrina Baroni (Liliaceae) is a perennial herb with antidepressant, antioxidant, antitumor, anti-inflammatory, and neuroprotective effects.The aim of our study was to investigate the effects of H. citrina extract (HCE) on SD-induced cognitive decline and depression-like behavior and possible neuroinflammation-related mechanisms. HCE (2 g/kg/day, i.g.) or vortioxetine (10 mg/kg/day, i.g.) were given to mice by oral gavage for a total of 28 days during the SD process. HCE treatment was found to ameliorate SD-induced impairment of short- and long-term spatial and nonspatial memory, measured using Y-maze, object recognition, and Morris water maze tests, as well as mitigating SD-induced depression-like behaviors, measured by tail suspension and forced swimming tests. HCE also reduced the levels of inflammatory cytokines (IL-1ß, IL-18, and IL-6) in the serum and hippocampus. Furthermore, HCE suppressed SD-induced microglial activation in the prefrontal cortex (PFC) and the CA1 and dentate gyrus (DG) regions of the hippocampus. HCE also inhibited the expression of phosphorylated NF-κB and activation of the NLRP3 inflammasome. In summary, our findings indicated that HCE attenuated SD-induced cognitive impairment and depression-like behavior and that this effect may be mediated by the inhibition of inflammatory progression and microglial activation in the hippocampus, as well as the down-regulation of NF-κB and NLRP3 signaling. The findings of these studies showingTthese results indicate that HCE exerts neuroprotective effects and are consistent with the findings of previous studies, suggesting that HCE is beneficial for the prevention and treatment of cognitive decline and depression in SD.

Molecular mechanism of Wilms tumour 1-associated protein in diabetes-related dry eye disease by mediating m6A methylation modification of lncRNA NEAT1.

Background: Dry eye disease (DED) is often secondary to diabetes mellitus (DM).Purpose: This study is to explore the action of Wilms tumor 1-associated protein (WTAP) in DM-DED via lncRNA NEAT1 m6A methylation.Methods: DM-DED mouse models were treated with sh-WTAP/sh-NEAT1, followed by assessment of corneal epithelial damage/histopathological changes. HCE-2 cells were exposed to hyperosmotic conditions to establish in vitro DED models and treated with oe-NEAT1/sh-NEAT1/sh-WTAP/nigericin (an NLRP3 inflammasome inducer). Cell viability/apoptosis were evaluated by CCK-8/TUNEL. Levels of WTAP/NEAT1/inflammatory factors/NLRP3 inflammasome- and apoptosis-related markers were determined. m6A modification was examined by MeRIP-qPCR and NEAT1 stability was also detected.Results: DM-DED mice exhibited up-regulated WTAP/NEAT1 expression and severe corneal damage, whereas WTAP/NEAT1 knockdown alleviated inflammation/corneal damage. In hyperosmolarity-induced HCE-2 cells, NEAT1 aggravated inflammation and apoptosis, while NEAT1 knockdown suppressed NLRP3 inflammasome activation and ameliorated cell injury. Hyperosmolarity-induced WTAP expression increased m6A modification and NEAT1 mRNA stability. WTAP mediated m6A methylation of NEAT1 and NLRP3 inflammasome activation in DM-DED mice.

Psychosocial stress induction in vivo vs. in virtuo and the influence of a health app on the acute stress reaction in youths: a study protocol for a randomized controlled trial.

BACKGROUND: Acute and everyday stress is substantial for the development of mental and physical diseases, therefore it is crucial to get a better understanding of its pathogenesis. Different methods (e.g., Ambulatory Assessment) and stress reactivity paradigms (e.g., Trier Social Stress Test / TSST) in laboratory settings are often used to investigate basic mechanisms of this process. Due to the technological progress of the last years and especially due to children and adolescents growing up with it, the application of these developments in clinical research is reasonable. The aim of this project is to successfully transfer the TSST for children and adolescents into the virtual world, which will be compared to a real TSST situation. Physiological and psychological stress reactions will be analyzed in order to assess similarities and differences. Moreover, it will be investigated whether a Heart Coherence Exercise (HCE) has a stronger influence on coping with acute stress compared to Natural Relaxation (NR). METHODS: This single-center experimental study will examine acute and everyday stress and coping processes in eighty-four healthy children and adolescents between the ages of 11 and 17. For everyday stress, different parameters (e.g., hormonal profiles and mood ratings) as well as a history of stressful life events and utilized coping methods will be recorded and a relaxation exercise will be practiced on a smartphone over 2 days. Regarding the acute stress reaction, the participants will be confronted either with the virtual or the real version of the TSST, followed by the trained relaxation exercise (HCE vs. NR). Physiological (e.g., cortisol and heart rate) and psychological stress markers (e.g., mood and gaze behavior) will be recorded continuously. DISCUSSION: Studies are sparse using a virtual version of the TSST in children and adolescents. A successful virtual TSST would constitute an economical variant, which would also make it easier to administer it in clinical or population-based samples. Effective ambulatory relaxation exercises would be a useful addition to clinical treatment approaches. TRIAL REGISTRATION: The study is registered in the German Clinical Trials Register since 10 August 2020 ( DRKS00022063 ).

Cellular localization of dieldrin and structure-activity relationship of dieldrin analogues in dopaminergic cells.

The incidence of Parkinson's disease (PD) correlates with environmental exposure to pesticides, such as the organochlorine insecticide, dieldrin. Previous studies found an increased concentration of the pesticide in the striatal region of the brains of PD patients and also that dieldrin adversely affects cellular processes associated with PD. These processes include mitochondrial function and reactive oxygen species production. However, the mechanism and specific cellular targets responsible for dieldrin-mediated cellular dysfunction and the structural components of dieldrin contributing to its toxicity (toxicophore) have not been fully defined. In order to identify the toxicophore of dieldrin, a structure-activity approach was used, with the toxicity profiles of numerous analogues of dieldrin (including aldrin, endrin, and cis-aldrin diol) assessed in PC6-3 cells. The MTT and lactate dehydrogenase (LDH) assays were used to monitor cell viability and membrane permeability after treatment with each compound. Cellular assays monitoring ROS production and extracellular dopamine metabolite levels were also used. Structure and stereochemistry for dieldrin were found to be very important for toxicity and other end points measured. Small changes in structure for dieldrin (e.g., comparison to the stereoisomer endrin) yielded significant differences in toxicity. Interestingly, the cis-diol metabolite of dieldrin was found to be significantly more toxic than the parent compound. Disruption of dopamine catabolism yielded elevated levels of the neurotoxin, 3,4-dihydroxyphenylacetaldehyde, for many organochlorines. Comparisons of the toxicity profiles for each dieldrin analogue indicated a structure-specific effect important for elucidating the mechanisms of dieldrin neurotoxicity.

Composition of aldrin, dieldrin, and photodieldrin enantiomers in technical and environmental samples.

Aldrin and dieldrin belong to the group of polycyclic chlorinated insecticides that are banned under the Stockholm Convention (POP Convention). Despite the fact that the use of these compounds ceased many years ago, aldrin and, in particular, dieldrin are still present in the environment from former applications, leading occasionally to contamination of agricultural produce and food, particularly Cucurbitaceae. These prochiral compounds have a complex stereochemistry. In the environment, aldrin is rapidly converted to its epoxide, dieldrin. Photolysis is one of the environmental transformation processes reported to be important for the compounds, leading to photoproducts such as photoaldrin and photodieldrin. In contrast to the parent compounds, photoaldrin and photodieldrin are chiral and exist as pairs of enantiomers. Although dieldrin and its metabolites have been extensively reviewed, the chirality of many of its metabolites has so far not been considered. In this study, the composition of technical aldrin and dieldrin from the 1950s and their photoproducts was investigated using both non-enantioselective and enantioselective gas chromatography with detection by several mass spectrometric techniques. Full enantiomer resolution of photodieldrin was achieved using a column with a silylated gamma-cyclodextrin as chiral selector. Photoaldrin, however, showed peak broadening, indicating some marginal resolution of the enantiomers. Whereas photodieldrin was formed as a racemate from both aldrin and dieldrin by natural sunlight, the analysis of environmental and biological samples (soil, biota) indicated its presence mostly with enantiomer compositions clearly differing from 1:1. The presence of photodieldrin in soil, treated more than 40 years ago with aldrin or dieldrin, documents that the photoreaction of dieldrin plays some role in the transformation of the compounds in the environment and that enantioselective biological processes are involved in its further transformation. The preliminary data also indicate that photodieldrin probably is not bioaccumulated more than dieldrin.

Effects of photoisomers of cyclodiene insecticides on liver and microsomal cytochrome P450 in rats.

Threshold dosages of the photoisomers of cyclodiene insecticides, namely photochlordane, photodieldrin, and photoheptachlor, for the induction of hepatic microsomal cytochrome P450 (P450) and liver hypertrophy in male rats were at least one-quarter of those reported for corresponding parent cyclodienes. Maximum increase in total P450 concentration (30%) and demethylases activities (100%) was always respectively one-third or one-tenth of that reported for parent cyclodienes. The P450 isozymic form induced by photoheptachlor resembled that induced by pentobarbital (P4502B1) in its substrate specificity, spectral characteristics, and electrophoretic mobility. The induction of P450 was initially followed by hepatic hypertrophy. However, higher dosages of photoisomers caused wasting and lowered both the liver weight and the activity of aniline hydroxylase while those of mirex and endrin, which also caused wasting and lowered aniline hydroxylase activity, continued causing further hepatic hypertrophy.

[Contamination of game in Schleswig-Holstein. 2. Residues of dieldrin, heptachlor epoxide and other cyclodiene insecticides in the liver fat of hares (Lepus europaeus L.)]. / Schadstoffbelastung von Wild aus Schleswig-Holstein. 2. Mitteilung: Rückstände von Dieldrin, Heptachlorepoxid und anderen Cyclodien-Insecticiden im Leberfett von Hasen (Lepus europaeus L.).

Increased levels of Dieldrin (0.01-2.94 mg/kg fat, median mean = 0.38 mg/kg fat) and cis-Heptachlor Epoxide (less than 0.01-1.38 mg/kg fat, mean = 0.20 mg/kg fat) were determined by GC/ECD in liver fat of 199 hares (Lepus europaeus L.) from Schleswig-Holstein, North Germany. This investigation was carried out within the scope of a game investigation program. The dependence of these residues on endogenous and exogenous factors are discussed. Oxychlordane and trans-Nonachlor, residues of Chlordane, were also analysed in liver fat by GC/MS. The application of the cyclodiene insecticides Aldrin/Dieldrin, Heptachlor and Chlordane has been prohibited in West Germany for several years; the residues indicate the high soil persistence of these substances.

Hepatic metabolism of cyclodiene insecticides by constitutive forms of cytochrome P-450 from lower vertebrates.

1. Multiple forms of cytochrome P-450 were separated from the hepatic microsomes of untreated male rats, pigeons (Columbia livia), razorbills (Alca torda), puffins (Fratercula arctica), and rainbow trout (Salmo gairdnerii), using anion exchange chromatography and DEAE-cellulose. 2. In some cases cytochrome P-450 forms were further purified on hydroxylapatite and carboxymethyl-sephadex columns. 3. Considerable differences in the distribution of forms between these five species were evident from elution profiles on DEAE cellulose, and on analysis of the cytochrome P-450 containing pools by SDS-PAGE. 4. The metabolism of two organochlorine compounds, aldrin and the dieldrin analogue HCE, were studied in (a) intact microsomes and (b) reconstituted systems containing cytochrome P-450, from each of the five species. 5. In spite of their close structural similarity, significant differences were found between the two substrates in the distribution of catalytic activity between the cytochrome P-450 isozymes of each species.

Dieldrin transformation to photodieldrin in liver and kidney of broiler.

Dieldrin, a potent residual insecticide and common contaminant of the environment, is metabolized in liver and kidney of broiler to photodieldrin, a photoisomer of dieldrin. This metabolite has by gas-liquid chromatography (g.l.c.) analysis a retention time different to the aldrin. The analysis by thin-layer chromatography (t.l.c.) of metabolite shows an Rf of 0.37. The i.r. (infra-red) spectrum shows the band corresponding to the epoxide ring (800-1300 cm-1) and does not show the characteristics bands of C = O (1800 cm-1) and ClC = CCl (1600 cm-1).

The hydration of sterically hindered epoxides by epoxide hydrolase of the rat and rabbit.

The hydration of the sterically-hindered epoxides dieldrin (HEOD) (1,2,3,4, 10,10-hexachloro-1,4,4a,5,6,7,8,8a-octahydro-6,7-epoxy-exo-1,4-endo-5,8- dimethanonaphthalene) and MME (1,2,3,4,9,9-hexachloro-1,4,4a,5,8,8a-hexahydro-6-methyl-6,7 epoxy 1,4-methanonaphthalene) was studied in pig and rabbit liver microsomes, and in apparently homogeneous preparations of epoxide hydrolase purified from liver microsomes of the rat and rabbit. A non-hindered substrate, HEOM (1,2,3,4,9,9-hexachloro-1,4,4a,5,6,7,8,8a-octahydro exo-6,7 epoxy-1,4 methano naphthalene) was used to assay both the enzyme preparations and the microsomes for epoxide hydrolase activity. The purified enzymes had more stable activity when incorporated into suspensions of phospholipid derived from rat liver microsomes than when used alone, so this preparation was used for long-term incubation with HEOD and MME. Activity towards the three substrates followed the sequence HEOM much greater than MME much much greater than HEOD, in the approximate ratios 1.5 X 10(6):10(4) activity towards dieldrin being only 0.0065-0.052 pmol/mg prot/min. The sole product of MME hydration was identified as a trans diol. The only product identified in the case of HEOD hydration by microsomes or enzyme preparations was the trans diol so there was no evidence for a significant metabolic pathway through the cis diol which is mediated by epoxide hydralase.

Effects of neonatally administered chlorpromazine and reserpine on the responsiveness of rat hepatic drug-metabolising enzymes to testosterone in adult life.

The effects of neonatally administered chlorpromazine and reserpine on the response of rat hepatic drug-metabolising enzymes to testosterone in adult life have been investigated using the chlorinated cyclodiene substrate DME. Neonatal treatment with chlorpromazine and reserpine had effects on the metabolism of DME similar to, but not as pronounced as, those of castration when adult. The effects of adult castration of male rats on hepatic microsomal metabolism of DME were fully reversed by treatment with testosterone propionate, with metabolism being restored to that of a control intact male. However, testosterone propionate treatment of either intact or castrated adult males that had received neonatal reserpine or chlorpromazine did not restore levels of metabolism to those characteristic of control adult male rats. These results suggest that neonatally administered chlorpromazine and reserpine alter the sensitivity of hepatic drug-metabolising enzymes to the actions of testosterone in adult life.